Evidences provided from several laboratories using genetically engineered mice indicated that activation of NF-kB is a key event associated with intestinal injury and bacterial host responses, and likely contributed to the maintenance of intestinal homeostasis. Upstream of the NF-kB transcriptional system lies two important signaling molecules involved in bacteria sensing/detection;MyD88 and Nod2. Although both proteins are critical for transmitting bacteria-induced NF-kB signaling, the intestine of either MyD88 or Nod2 gene deficient mice are normal with no sign of intestinal inflammation or impaired function in a normal environment (pathogen free). While there is a reasonably good understanding on how bacteria trigger NF-kB signaling in the intestine, it is largely unclear how this event leads to either cytoprotective or proinflammatory responses. Lack of understanding on how bacteria induced protective or deleterious response through NF-kB signaling undermines the establishment of therapeutic strategies aimed at alleviating inflammatory disorders. The central hypothesis for the proposed project is that MyD88 and Nod2-derived NF-kB signaling differentially impact on the state of intestinal response to the microbiota and consequently intestinal inflammation. We have formulated this hypothesis based on our findings showing that disrupting MyD88 signaling prevented the development of spontaneous colitis in IL-10-/- mice (IL-10-/-;MyD88-/-), whereas disrupting Nod2 signaling (IL-10-/-;Nod2-/-) exacerbate colitis in the same model. In addition, our temporal and spacial analysis of NF-kB activity using gnotobiotic IL-10-/-;NF-kBEGFP mice show a rapid and transient EGFP activation in IEC but a sustain expression in the intestinal lamina propria immune cells following bacteria colonization. We plan to test our central hypothesis and fulfill the overall objective of this application with the following specific aims. AIM #l, Define the physiological role of Nod2/NF-kB signaling in regulating bacteria-mediated intestinal inflammation. The working hypothesis for this aim, based on preliminary data is that Nod2-dependent NF-kB signaling help maintain intestinal homeostasis through induction of protective molecules and negative feedback to toll-like receptor, which then prevent dysregulated host response to the microbiota. AIM #2 Dissect the contribution of IEC- and myeloid-derived MyD88/NF-:B signaling in bacteria-mediated intestinal inflammation. The working hypothesis for this aim is that production of IEC- and myeloid-derived NF-B mediators differentially impact on the state of intestinal response to the microbiota and consequently inflammation. The predominant role of IEC-derived NF-kB signaling is likely to protect the integrity of the epithelium through production of anti-apoptotic and anti-microbial genes. As opposed, myeloid-derived NF-kB signaling likely drives expression of pro- inflammatory genes implicated in the innate/adaptive host response to microorganisms. At the completion of these studies, our expectation is that we will have determined the mechanisms by which MyD88 and Nod2-derived NF-kB signaling impact on bacteria/host responses and intestinal inflammation. Since dysregulated host response to the intestinal microbiota associate with the development of both ulcerative colitis and Crohn's disease, our findings will provide novel means to prevent/treat this pathological conditions. PUBLIC HEALTH RELEVANCE: Inflammatory bowel diseases (IBD) such as ulcerative colitis (UC) and Crohn's disease represent a major health problem in North America, with over 1.4 million people suffering from one of these two diseases. In the past decade, the intestinal microbiota has taken center stage in the etiology of IBD. The signaling protein MyD88 and Nod2 are important factor involved in bacteria-mediated host responses. This project investigates how MyD88 and Nod2 signaling to the transcription factor NF-kB control intestinal homeostasis and disease development. Since dysregulated host response to the intestinal microbiota associate with the development of both ulcerative colitis and Crohn's disease, our study will potentially identify novel means to prevent/treat IBD.